The genetic and clinical spectrum in a cohort of 39 families with complex inherited peripheral neuropathies.

With complicated conditions and a large number of potentially causative genes, the diagnosis of a patient with complex inherited peripheral neuropathies (IPNs) is challenging. To provide an overview of the genetic and clinical features of 39 families with complex IPNs from central south China and to optimize the molecular diagnosis approach to this group of heterogeneous diseases, a total of 39 index patients from unrelated families were enrolled, and detailed clinical data were collected. TTR Sanger sequencing, hereditary spastic paraplegia (HSP) gene panel, and dynamic mutation detection in spinocerebellar ataxia (SCAs) were performed according to the respective additional clinical features. Whole-exome sequencing (WES) was used in patients with negative or unclear results. Dynamic mutation detection in NOTCH2NLC and RCF1 was applied as a supplement to WES. As a result, an overall molecular diagnosis rate of 89.7% was achieved. All 21 patients with predominant autonomic dysfunction and multiple organ system involvement carried pathogenic variants in TTR, among which nine had c.349G > T (p.A97S) hotspot variants. Five out of 7 patients (71.4%) with muscle involvement harbored biallelic pathogenic variants in GNE. Five out of 6 patients (83.3%) with spasticity reached definite genetic causes in SACS, KIF5A, BSCL2, and KIAA0196, respectively. NOTCH2NLC GGC repeat expansions were identified in all three cases accompanied by chronic coughing and in one patient accompanied by cognitive impairment. The pathogenic variants, p.F284S and p.G111R in GNE, and p.K4326E in SACS, were first reported. In conclusion, transthyretin amyloidosis with polyneuropathy (ATTR-PN), GNE myopathy, and neuronal intranuclear inclusion disease (NIID) were the most common genotypes in this cohort of complex IPNs. NOTCH2NLC dynamic mutation testing should be added to the molecular diagnostic workflow. We expanded the genetic and related clinical spectrum of GNE myopathy and ARSACS by reporting novel variants.

Comprehensive Genetic Analyses of Inherited Peripheral Neuropathies in Japan: Making Early Diagnosis Possible.

Various genomic variants were linked to inherited peripheral neuropathies (IPNs), including large duplication/deletion and repeat expansion, making genetic diagnosis challenging. This large case series aimed to identify the genetic characteristics of Japanese patients with IPNs. We collected data on 2695 IPN cases throughout Japan, in which PMP22 copy number variation (CNV) was pre-excluded. Genetic analyses were performed using DNA microarrays, next-generation sequencing-based gene panel sequencing, whole-exome sequencing, CNV analysis, and RFC1 repeat expansion analysis. The overall diagnostic rate and the genetic spectrum of patients were summarized. We identified 909 cases with suspected IPNs, pathogenic or likely pathogenic variants. The most common causative genes were MFN2, GJB1, MPZ, and MME. MFN2 was the most common cause for early-onset patients, whereas GJB1 and MPZ were the leading causes of middle-onset and late-onset patients, respectively. Meanwhile, GJB1 and MFN2 were leading causes for demyelinating and axonal subtypes, respectively. Additionally, we identified CNVs in MPZ and GJB1 genes and RFC1 repeat expansions. Comprehensive genetic analyses explicitly demonstrated the genetic basis of our IPN case series. A further understanding of the clinical characteristics of IPN and genetic spectrum would assist in developing efficient genetic testing strategies and facilitate early diagnosis.

Nerve pathology in animal models of neuropathies.

To understand the pathology of axonal degeneration and demyelination in peripheral neuropathy, histological investigations in different animal models that mimic some aspects of human peripheral neuropathy are needed. Thus, in the following section of this special issue, the main pathological features of experimental autoimmune neuritis, animal models of chemotherapy-induced peripheral neuropath and of human inherited peripheral neuropathies (IPNs) will be illustrated. When possible, micrographs from animal models and selected human biopsy will be shown side by side.

Targeted Therapies for Hereditary Peripheral Neuropathies: Systematic Review and Steps Towards a 'treatabolome'.

BACKGROUND: Hereditary peripheral neuropathies are inherited disorders affecting the peripheral nervous system, including Charcot-Marie-Tooth disease, familial amyloid polyneuropathy and hereditary sensory and motor neuropathies. While the molecular basis of hereditary peripheral neuropathies has been extensively researched, interventional trials of pharmacological therapies are lacking. OBJECTIVE: We collated evidence for the effectiveness of pharmacological and gene-based treatments for hereditary peripheral neuropathies. METHODS: We searched several databases for randomised controlled trials (RCT), observational studies and case reports of therapies in hereditary peripheral neuropathies. Two investigators extracted and analysed the data independently, assessing study quality using the Oxford Centre for Evidence Based Medicine 2011 Levels of Evidence in conjunction with the Jadad scale. RESULTS: Of the 2046 studies initially identified, 119 trials met our inclusion criteria, of which only 34 were carried over into our final analysis. Ascorbic acid was shown to have no therapeutic benefit in CMT1A, while a combination of baclofen, naltrexone and sorbitol (PXT3003) demonstrated some efficacy, but phase III data are incomplete. In TTR-related amyloid polyneuropathy tafamidis, patisiran, inotersen and revusiran showed significant benefit in high quality RCTs. Smaller studies showed the efficacy of L-serine for SPTLC1-related hereditary sensory neuropathy, riboflavin for Brown-Vialetto-Van Laere syndrome (SLC52A2/3) and phytanic acid-poor diet in Refsum disease (PHYH). CONCLUSIONS: The 'treatable' variants highlighted in this project will be flagged in the treatabolome database to alert clinicians at the time of the diagnosis and enable timely treatment of patients with hereditary peripheral neuropathies.

Analysis of the conformational changes caused by the mutations in mitofusin2 gene by Insilico approach.

OBJECTIVE: To find the effect of pathogenic Mitofusin 2 mutations, responsible for Charcot-Marie-Tooth hereditary neuropathy type 2A, on protein structure. METHODS: The study was conducted at department of biosciences COMSATS University Islamabad, Sahiwal campus from September 2016 to July 2017, and comprised patients with Charcot Marie-Tooth hereditary neuropathy type 2A who were divided into early-onset severe group A and late-onset mild group B. Bioinformatics and molecular analysis was done to find the changes in the protein structure caused by the mutation. Three mutations were selected in two domains of the gene. These were: p. Arg94Trp, p. His165Arg and p. Thr362Met. RESULTS: Of the 10 patients, 5(50%) were in each of the two groups. Change in the structure was predicted in the mutated protein at position p. Arg94Trp, and, due to the mutation, an extra alpha helix was formed in the mutated protein. CONCLUSIONS: Change in the structure of protein can be in a critical position that is involved in the mitochondrial fusion process. However, further studies are required to validate and explain the findings.

Defects in Axonal Transport in Inherited Neuropathies.

Axonal transport is a highly complex process essential for sustaining proper neuronal functioning. Disturbances can result in an altered neuronal homeostasis, aggregation of cargoes, and ultimately a dying-back degeneration of neurons. The impact of dysfunction in axonal transport is shown by genetic defects in key proteins causing a broad spectrum of neurodegenerative diseases, including inherited peripheral neuropathies. In this review, we provide an overview of the cytoskeletal components, molecular motors and adaptor proteins involved in axonal transport mechanisms and their implication in neuronal functioning. In addition, we discuss the involvement of axonal transport dysfunction in neurodegenerative diseases with a particular focus on inherited peripheral neuropathies. Lastly, we address some recent scientific advances most notably in therapeutic strategies employed in the area of axonal transport, patient-derived iPSC models, in vivo animal models, antisense-oligonucleotide treatments, and novel chemical compounds.

Molecular diagnosis of inherited peripheral neuropathies by targeted next-generation sequencing: molecular spectrum delineation.

PURPOSE: Inherited peripheral neuropathies (IPN) represent a large heterogenous group of hereditary diseases with more than 100 causative genes reported to date. In this context, targeted next-generation sequencing (NGS) offers the opportunity to screen all these genes with high efficiency in order to unravel the genetic basis of the disease. Here, we compare the diagnostic yield of targeted NGS with our previous gene by gene Sanger sequencing strategy. We also describe several novel likely pathogenic variants. DESIGN AND PARTICIPANTS: We have completed the targeted NGS of 81 IPN genes in a cohort of 123 unrelated patients affected with diverse forms of IPNs, mostly Charcot-Marie-Tooth disease (CMT): 23% CMT1, 52% CMT2, 9% distal hereditary motor neuropathy, 7% hereditary sensory and autonomic neuropathy and 6.5% intermediate CMT. RESULTS: We have solved the molecular diagnosis in 49 of 123 patients (~40%). Among the identified variants, 26 variants were already reported in the literature. In our cohort, the most frequently mutated genes are respectively: MFN2, SH3TC2, GDAP1, NEFL, GAN, KIF5A and AARS. Panel-based NGS was more efficient in familial cases than in sporadic cases (diagnostic yield 49%vs19%, respectively). NGS-based search for copy number variations, allowed the identification of three duplications in three patients and raised the diagnostic yield to 41%. This yield is two times higher than the one obtained previously by gene Sanger sequencing screening. The impact of panel-based NGS screening is particularly important for demyelinating CMT (CMT1) subtypes, for which the success rate reached 87% (36% only for axonal CMT2). CONCLUSION: NGS allowed to identify causal mutations in a shorter and cost-effective time. Actually, targeted NGS is a well-suited strategy for efficient molecular diagnosis of IPNs. However, NGS leads to the identification of numerous variants of unknown significance, which interpretation requires interdisciplinary collaborations between molecular geneticists, clinicians and (neuro)pathologists.

A Review of Copy Number Variants in Inherited Neuropathies.

The rapid development in the last 10-15 years of microarray technologies, such as oligonucleotide array Comparative Genomic Hybridization (CGH) and Single Nucleotide Polymorphisms (SNP) genotyping array, has improved the identification of fine chromosomal structural variants, ranging in length from kilobases (kb) to megabases (Mb), as an important cause of genetic differences among healthy individuals and also as disease-susceptibility and/or disease-causing factors. Structural genomic variations due to unbalanced chromosomal rearrangements are known as Copy-Number Variants (CNVs) and these include variably sized deletions, duplications, triplications and translocations. CNVs can significantly contribute to human diseases and rearrangements in several dosage-sensitive genes have been identified as an important causative mechanism in the molecular aetiology of Charcot-Marie-Tooth (CMT) disease and of several CMT-related disorders, a group of inherited neuropathies with a broad range of clinical phenotypes, inheritance patterns and causative genes. Duplications or deletions of the dosage-sensitive gene PMP22 mapped to chromosome 17p12 represent the most frequent causes of CMT type 1A and Hereditary Neuropathy with liability to Pressure Palsies (HNPP), respectively. Additionally, CNVs have been identified in patients with other CMT types (e.g., CMT1X, CMT1B, CMT4D) and different hereditary poly- (e.g., giant axonal neuropathy) and focal- (e.g., hereditary neuralgic amyotrophy) neuropathies, supporting the notion of hereditary peripheral nerve diseases as possible genomic disorders and making crucial the identification of fine chromosomal rearrangements in the molecular assessment of such patients. Notably, the application of advanced computational tools in the analysis of Next-Generation Sequencing (NGS) data has emerged in recent years as a powerful technique for identifying a genome-wide scale complex structural variants (e.g., as the ones resulted from balanced rearrangements) and also smaller pathogenic (intragenic) CNVs that often remain beyond the detection limit of most conventional genomic microarray analyses; in the context of inherited neuropathies where more than 70 disease-causing genes have been identified to date, NGS and particularly Whole-Genome Sequencing (WGS) hold the potential to reduce the number of genomic assays required per patient to reach a diagnosis, analyzing with a single test all the Single Nucleotide Variants (SNVs) and CNVs in the genes possibly implicated in this heterogeneous group of disorders.

HERC1 Ubiquitin Ligase Is Required for Normal Axonal Myelination in the Peripheral Nervous System.

A missense mutation in HERC1 provokes loss of cerebellar Purkinje cells, tremor, and unstable gait in tambaleante (tbl) mice. Recently, we have shown that before cerebellar degeneration takes place, the tbl mouse suffers from a reduction in the number of vesicles available for release at the neuromuscular junction (NMJ). The aim of the present work was to study to which extent the alteration in HERC1 may affect other cells in the nervous system and how this may influence the motor dysfunction observed in these mice. The functional analysis showed a consistent delay in the propagation of the action potential in mutant mice in comparison with control littermates. Morphological analyses of glial cells in motor axons revealed signs of compact myelin damage as tomacula and local hypermyelination foci. Moreover, we observed an alteration in non-myelinated terminal Schwann cells at the level of the NMJ. Additionally, we found a significant increment of phosphorylated Akt-2 in the sciatic nerve. Based on these findings, we propose a molecular model that could explain how mutated HERC1 in tbl mice affects the myelination process in the peripheral nervous system. Finally, since the myelin abnormalities found in tbl mice are histological hallmarks of neuropathic periphery diseases, tbl mutant mice could be considered as a new mouse model for this type of diseases.

Structural variations causing inherited peripheral neuropathies: A paradigm for understanding genomic organization, chromatin interactions, and gene dysregulation.

Inherited peripheral neuropathies (IPNs) are a clinically and genetically heterogeneous group of diseases affecting the motor and sensory peripheral nerves. IPNs have benefited from gene discovery and genetic diagnosis using next-generation sequencing with over 80 causative genes available for testing. Despite this success, up to 50% of cases remain genetically unsolved. In the absence of protein coding mutations, noncoding DNA or structural variation (SV) mutations are a possible explanation. The most common IPN, Charcot-Marie-Tooth neuropathy type 1A (CMT1A), is caused by a 1.5 Mb duplication causing trisomy of the dosage sensitive gene PMP22. Using genome sequencing, we recently identified two large genomic rearrangements causing IPN subtypes X-linked CMT (CMTX3) and distal hereditary motor neuropathy (DHMN1), thereby expanding the spectrum of SV mutations causing IPN. Understanding how newly discovered SVs can cause IPN may serve as a useful paradigm to examine the role of topologically associated domains (TADs), chromatin interactions, and gene dysregulation in disease. This review will describe the growing role of SV in the pathogenesis of IPN and the importance of considering this type of mutation in Mendelian diseases where protein coding mutations cannot be identified.

A newly distal hereditary motor neuropathy caused by a rare AIFM1 mutation.

In two siblings, who suffer from an early childhood-onset axonal polyneuropathy with exclusive involvement of motor fibers, the c.629T>C (p.F210S) mutation was identified in the X-linked AIFM1 gene, which encodes for the apoptosis-inducing factor (AIF). The mutation was predicted as deleterious, according to in silico analysis. A decreased expression of the AIF protein, altered cellular morphology, and a fragmented mitochondrial network were observed in the proband's fibroblasts. This new form of motor neuropathy expands the phenotypic spectrum of AIFM1 mutations and therefore, the AIFM1 gene should be considered in the diagnosis of hereditary motor neuropathies.

Improving diagnosis of inherited peripheral neuropathies through gene panel analysis.

BACKGROUND: Inherited peripheral neuropathies (IPN) are the most common inherited neurological condition. It represents a highly heterogeneous group, both clinically and genetically. Targeted disease specific gene panel massively parallel sequencing (MPS) seems to be a useful tool in diagnosis of disorders with high genetic heterogeneity. METHODS: In our study, we have designed, validated and updated our own custom gene panel of all known genes associated with IPN. One hundred and ninety-eight patients have been tested so far. Only patients in whom mutations in more common causes or relevant genes have already been excluded were enrolled. Five consecutive panel designs were prepared according to recent literature search, the last one covering ninety-three genes. Each patient was tested only once. All data were evaluated with at least two different pipelines. RESULTS: In summary, causative mutation has been found in fifty-one patients (26 %). The results were inconclusive in thirty-one (16 %) patients. No variants of likely significance to IPN were found in one hundred and sixteen (58 %) patients. CONCLUSION: MPS gene panel enables testing of all known IPN causes at once with high coverage and at an affordable cost making it truly a method of choice also in IPN. Gene panel testing results in several interesting results and findings.